The conditioning of ethanol's (EtOH) reinforcing effects with environmental stimuli is a major factor in the abuse potential of this drug. EtOH-related stimuli elicit strong EtOH seeking in animal models of relapse and these models are widely employed to study the neurobiological basis of EtOH craving and relapse. However, behavioral and neurobiological information on conditioning factors in EtOH seeking from animal models is limited largely to that from EtOH nondependent subjects. In alcoholics, a significant positive correlation exists between the history of dependence and the severity of cue-induced EtOH craving. EtOH consumption during withdrawal modifies subjects' EtOH reinforcement history to include learning about amelioration or avoidance of adverse withdrawal states as a novel and essential aspect of EtOH's reinforcing actions, rendering the drug a qualitatively different, more potent reinforcer. Thus, understanding the control of behavior by stimuli conditioned to EtOH under conditions that encompass the reinforcing dimension of this drug that emerges with the experience of withdrawal states will be essential for advancing the understanding and treatment EtOH addiction. Preliminary data that stimuli conditioned to EtOH reinforcement during withdrawal elicit significant reinstatement confirm the need for this understanding. The purpose of this exploratory and developmental project is to investigate the significance of withdrawal-related conditioning in EtOH seeking and to gain understanding of the neurocircuitry regulating this behavior implementing novel methodology. Behavioral hypotheses to be tested are (a) that withdrawal-related conditioning overrides the control of EtOH seeking by cues conditioned to EtOH earlier in the nondependent state, (b) that, compared to cues conditioned to EtOH in the nondependent state, stimuli conditioned to EtOH reinforcement during withdrawal produce greater EtOH seeking following a stress challenge, and (c) greater resistance to conditioned suppression of EtOH seeking. Parallel studies will establish neural activation patterns associated with EtOH seeking in rats with and without histories of withdrawal-related learning using quantitative Fos immunohistochemistry. These studies are guided by the hypothesis that cues conditioned to EtOH during withdrawal produce a different pattern of neural activation, with a stronger engagement of stress-regulatory regions, compared to cues conditioned to EtOH in the nondependent state. Finally, to confirm a role of brain regions in EtOH seeking linked to withdrawal-related conditioning, Daun 02 pharmacogenetic inactivation in cfos-lacZ transgenic rats will be employed to selectively lesion neuronal ensembles in these sites. These developmental studies will test the hypothesis that neuronal ensembles can be identified within brain reward and stress circuitry that preferentially mediate the effects of stimuli conditioned to EtOH reinforcement during withdrawal. This research will provide the groundwork for future in-depth investigations of the significance of withdrawal-related conditioning in the abuse potential of EtOH to advance understanding of the neurobiology and of treatment strategies for alcoholism. PUBLIC HEALTH RELEVANCE: This proposal addresses the need for better understanding of the neurobehavioral basis of the compulsive and chronically relapsing nature of alcohol addiction. Neurobiological and medications development information from animal models of relapse to alcohol abuse is limited largely to studies in alcohol nondependent animals. To advance the understanding of alcohol addiction and improve treatment strategies, the proposed studies will investigate alcohol seeking induced by stimuli conditioned to the potent reinforcing dimensions of this drug that emerge with the experience of amelioration of withdrawal distress by EtOH consumption as well as the neurobiological basis of this behavior.